Synaptic and membrane changes have long been proposed to underlie learning and memory. Other than random sampling, it has been extremely difficult to characterize synaptic and membrane changes in individual, identified cells involved in learning. Rat eyeblink conditioning provides an opportunity to use transsynaptic tracers to identify which of the more than four thousand principal cells in the anterior interpositus receive tone and shock sensory inputs and provide the eyeblink motor output involved in this fundamental form of learning. We will take advantage of retrograde and anterograde transsynaptic tracers, immunohistochemistry in fixed tissue, and electrophysiological recordings in brain slices to identify and characterize synaptic and membrane properties of sensory input- and motor output-labeled neurons involved in rat eyeblink conditioning. A comparison of synaptic and membrane properties in cells from rats given tone-shock pairings with cells from rats given unpaired tone/shock presentations or simply handled and given chamber exposure, this approach will provide an understanding of how the synaptic and membrane properties of single cells identified to be at the nexus of an input-output circuit necessary for eyeblink conditioning are changed as a result of associative learning. We have a long history of eyeblink conditioning, have experience in both immunohistochemical identification of synaptic changes that result from learning and rat cerebellar slice whole-cell recordings, and have used both retrograde and anterograde transsynaptic tracing techniques to map the neural circuits involved in the eyeblink - all documenting the feasibility of this approach. We have compelling preliminary data documenting learning-specific increases in both excitatory and inhibitory synapses in labeled anterior interpositus neurons and data characterizing the intrinsic membrane properties of cells in rat deep cerebellar nuclei identified by a motor output transsynaptic tracer. The challenge - and reason for a high-risk developmental application - is to now combine these different research components into a single set of experiments to characterize the synaptic and membrane changes in individual, identified cells as a result of eyeblink conditioning in an essential learning circuit.